Intracellular localization of human DNA polymerase alpha with monoclonal antibodies.
Intracellular localization of human DNA polymerase alpha with monoclonal antibodies.
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DOI：
10.1016/s0021-9258(18)34344-8
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发表时间：
1982-07
期刊：
影响因子：
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通讯作者：
K. Bensch;Shoichiro Tanaka;S. Z. Hu;T. S. Wang;D. Korn
中科院分区：
文献类型：
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作者：
K. Bensch;Shoichiro Tanaka;S. Z. Hu;T. S. Wang;D. Korn
我们已经成功地建立了16个稳定的小鼠杂交瘤，分泌特异性的人DNA聚合酶α的单克隆抗体。免疫细胞化学研究的结果，使用4这些单克隆抗体和免疫过氧化物酶检测方法，文件的DNA聚合酶α在三个单独的线培养的人细胞的专门的核内定位。通过光学显微镜，免疫过氧化物酶反应产物在核质内呈弥散分布，但核仁明显呈阴性。在转化系KB和BeWo的培养物中，超过955个细胞呈阳性，表明核内DNA聚合酶α抗原在整个有丝分裂周期中持续存在。与此形成鲜明对比的是，在正常二倍体成纤维细胞系WI-38中，培养细胞的较小部分是阳性的，并且在推测为接触停滞且不再有丝分裂循环的小菌落的紧密贴壁细胞中没有可检测到的聚合酶α抗原。在有丝分裂的细胞中，核膜溶解（因此暂时无核），抗聚合酶α反应继续呈强阳性，在这种情况下，反应产物弥漫分布在整个细胞质中。KB细胞的初步电子显微镜检查证实并扩展了这些观察结果。免疫过氧化物酶反应产物基本上限于核室，主要分布在核质的中间区域，位于中央的核仁和位于外周的浓缩染色质块之间。
We have successfully established 16 stable murine hybridomas that secrete monoclonal antibodies specific for human DNA polymerase alpha. The results of immunocytochemical studies, using 4 of these monoclonal antibodies and immunoperoxidase detection methods, document the exclusively intranuclear localization of DNA polymerase alpha in three separate lines of cultured human cells. By light microscopy, the immunoperoxidase reaction product exhibits a diffuse pattern of distribution within the nucleoplasm, but nucleoli are clearly negative. In cultures of the transformed lines, KB nd BeWo, more than 955 of the cells are positive, suggesting that intranuclear DNA polymerase alpha antigens persist throughout the mitotic cycle. In striking contrast, in the normal diploid fibroblast line, WI-38, a smaller fraction of the cultured cells is positive, and there is no detectable polymerase alpha antigen in the closely apposed cells of microcolonies that are presumed to be contact-arrested and no longer mitotically cycling. In cells in mitosis that have dissolved their nuclear envelopes (and are thus transiently anucleate), the anti-polymerase alpha reaction continues to be strongly positive, and in this single circumstance the reaction product is diffusely distributed throughout the cell cytoplasm. Initial electron microscopic examination of KB cells confirms and extends these observations. The immunoperoxidase reaction product is essentially limited to the nuclear compartment and is predominantly distributed in the midzonal region of the nucleoplasm between centrally disposed nucleoli and peripherally located blocks of condensed chromatin.